The major goal of this research is to delineate mechanisms whereby nutritional factors and pharmacological agents influence the metabolic activation and conjugation of precarcinogens and model drug substrates in intact cells using isolated perfused organs as experimental models. The following studies are proposed: (1) to investigate the effect of NADPH supply on rates of mixed-function oxidation in the isolated hemoglobin-free perfused rat liver and rabbit lung using benzo(a)pyrene, 7-ethoxycoumarin and p-nitroanisole as substrates. Alterations in mixed-function oxidation that occur in the presence of specific inhibitors, inducing agents and changes in nutritional status will be explored to determine the relative contribution of mitochondrial and cytosolic NADH and NADPH as regulators. Alterations is steady-state concentrations of key intermediates of energy metabolism, pyridine nucleotides, and oxidation-reduction induced changes in surface fluorescence of intracellular pigments will be correlated with changes in rates of mixed function oxidation. (2) to study the effect of altered nutritional status and selected inducing agents on rates of conjugation of the oxidized products of mixed-function oxidation in isolated perfused tissues. Rates of conjugation will be compared with specific enzyme activities and measured concentrations of activated intermediates such as UDPGA and PAPS. Information obtained in these studies will be applied to enhancing rates of conjugation of electrophiles generated from carcinogens in intact tissues. (3) Interactions between mixed-function oxidation, conjugation and intermediary metabolism will be studied in perfused livers of mice known to differ in their susceptibility to chemical carcinogens (e.g., DBA/2J and C57BL/6J). Also, the influence of nutritional factors and inducing agents on the microheterogeneity of oxygen gradients and enzymes associated with the formation of NADPH, intermediates of energy metabolism and rates of mixed-function oxidation will be examined in intact tissue via microfluorometry.